This invention relates to the field of capillary electrophoresis-based screening of materials for unidentified compounds that can bind to a target molecule of interest.
Developing screens to identify new, biologically active compounds can present unique and difficult challenges, especially when screening complex materials, particularly “complex biological materials”: any material that may have an effect in a biological system. Examples of complex material include, but are not limited to: naturally occurring complex biological materials, such as natural products or extracts; various biological preparations; chemical mixtures; libraries of pure compounds; and synthetic compounds such as combinatorial libraries. Examples of major screening problems include: detecting candidate hit compounds that bind to a target molecule of interest, especially those present at low concentrations in screened samples; accounting for unknown components that can interfere with screening agents; and determining the relative value of screened samples for further investigative efforts. As well, high concentrations of a weak or several weak, competing binder(s) can mask the signal from a moderate-to-tight-binding affinity ligand occurring at a lower concentration within the same sample.
Therefore, there remains a need for rapid and cost-effective screening tools for discovering new bioactive compounds and potential regulatory compounds, particularly those that bind to essential molecules of key metabolic pathways or molecules implicated in disease. Also needed is a way of prioritizing, for further characterization and testing, samples of material identified to contain potential candidate ligands and/or detected candidate ligands. The present invention addresses these needs, by providing a means of detecting unknown or unindentified ligands that may be candidate, new, bioactive compounds. In particular, the invention provides a means of better identifying ligands having a relative binding strength at or higher than a desired threshold as set by use of a known, preferably detectable, competitive ligand having a selected binding strength or dissociation constant (e.g., Kd≦100 μM, preferably Kd≦10 μM). Identifying and ranking those ligand-containing samples that form the most stable complexes with the selected target, saves time and resources spent on further isolation and characterization of hit compounds. The most stable ligands are potentially more effective and valuable as therapeutic, regulatory and/or diagnostic compounds and drugs.